2.Differentiation of adult adipose tissue-derived mesenchymal stem cells into cardiocyte-like cells in vitro
Guanghui CHEN ; Jing XIA ; Yuhong QIN
Chinese Journal of Tissue Engineering Research 2007;11(37):7494-7497
BACKGROUND: At present, many problems deserve to be solved before clinical utilization of adipose tissue-derived mesenchymal stem cells (ADMSCs) such as complete differentiation from ADMSCs into cardiomyocytes and ADMSCs with or without specific function of cardiomyocytes. It is of significance to solve the problems including how to elevate the differentiation rate of ADMSCs into cardiomyocytes and how to elevate the homing and survival rate after transplantation for clinical utilization of stem cells.OBJECTIVE: To observe the differentiation of ADMSCs into cardiomyocytes after in vitro culture and induction.DESIGN: Randomized controlled observation.SETTING: Laboratory of Cardiology, General Hospital of Chinese PLA.MATERIALS: Adipose tissue was collected from abdominal operative patients at Department of General Surgery of General Hospital of Chinese PLA with the agreement of patients. Iscove's Modified Dulbecco's Medium (IMDM), type Ⅰ collagenase, 5-azacytidine (5-aza) and polyclonal antibody of specific anti-myocardial Troponin T (TnT) were purchased from Hyclon company, Gibco company, Sigma and Fujian Maixin Biotechnology Company, respectively.Monoclone antibodies of CD44, CD45, CD34, HLA2DR and factor-Ⅷ, Desmin, anti-α-striated muscle actin and anti-myosin heavy chain (MHC) were bought from Beijing Zhongshan Golden Bridge Biotechnology Limited Corporation. DAB stain, reverse transcription-polymerase chain reaction (RT-PCR) kit and atrial natriuretic peptide (ANP) radioimmunity kit were purchased from Beijing Zhongshan Golden Bridge Biotechnology Limited Corporation,Invitrogen and Radioimmunity Research Institute of Science and Technology Development Center of General Hospital of Chinese PLA, respectively.METHODS: The experiment was performed at the Laboratory of Cardiology, General Hospital of Chinese PLA from March 2005 to April 2006. ADMSCs were isolated and cultured by digestion and attachment culture method. The third generation of cells were determined by immunocytochemical method for surface molecule CD44, CD13, CD105, CD45,CD34, HLA-DR, factor Ⅷ and induced by addition of 5-aza into culture medium. On the 7th, 14th, 21st and 28th days, gene of GATA4 and Nkx2.5 were tested by reverse transcription-polymerase chain reaction (RT-PCR), and the concentration of atrial natriuretic polypeptide (ANP) were also examined by radioimmunoassay.MAIN OUTCOME MEASURES: Determination of cell surface marker, gene of GATA4 and Nkx2.5, and ANP concentration.and factor Ⅷ negative for the cultured cells. After induction for 7 days with 5-aza, no cells expressed Desmin,α-Sarcomeric Actin, Myocin heavy chain (MHC) and TnT, most cells were positive stained for CD44. After induction for 14 days, small amounts of cells displayed positive for Desmin,α-Sarcomeric Actin and MHC but still negative for TnT,while partial cells expressed positive CD44. After induction for 21 days, most cells were positive for Desmin,α-Sarcomeric after induction and no significant difference was found compared with that at the 7th day [(0.022±0.01) ng/L (P>0.05].ANP concentration was respectively (7.92±0.21) and (8.12±0.50) ng/L at the 21st day and the 28th day (P>0.05), but there was significant difference compared with that at the 7th day (P<0.05).
3.Clinical diagnosis and treatment analysis of head and neck malignant tumor patients with postoperative pulmonary embolism
Jing ZHOU ; Xiaohong CHEN ; Mingzhao QIN
Chinese Archives of Otolaryngology-Head and Neck Surgery 2015;(10):504-506
[ABSTRACT]OBJECTIVE To summarize the clinical related factors and prognostic influence factors of perioperative pulmonary embolism of head and neck malignant tumor.METHODSFrom 2010-2014, our hospital carried out a total of 2736 cases of head and neck malignant tumor surgical operations, of which, 10 cases were clinically diagnosed as postoperative pulmonary embolism, retrospectively analyzed the process of clinical treatment of the patients of pulmonary embolism with head and neck malignant tumors, and summarized their etiological factors, clinical manifestations, diagnosis and treatment. RESULTS The preoperative period incidence of pulmonary embolism in patients with head and neck malignant tumor was 0.37%(10/2736). Their clinical manifestations were mainly of asthma and breathing difficulty, and 3 cases of asymptomatic hypoxemia. 8 cases of patients showed pulmonary artery and branch filling defects after pulmonary artery angiography (CTA), 7 cases of patients got cured and were discharged from the hospital after comprehensive treatment such as anticoagulation etc; 3 cases of patients died after emergency treatment. 2 cases of patients suffered cavity bleeding, and there was no anticoagulant drug adjustment.CONCLUSIONThe mortality of head and neck cancer patients with perioperative pulmonary embolism is high, and therefore, preventive measures and timely treatments are important to reduce the incidence of pulmonary embolism.
4.Effect of trypsin on the growth of a skin squamous cell carcinoma cell line A431
Jie ZHANG ; Yixin CAO ; Jing QIN ; Jianli WANG ; Li CHEN
Chinese Journal of Dermatology 2015;48(7):493-495
Objective To evaluate the effect of try psin on the proliferation,migration and adhesion of a skin squamous cell carcinoma cell line A431.Methods Cultured A431 cells were divided into several experimental groups treated with trypsin at concentrations of 0.1,1,10 and 100 nmol/L for 24,48 and 72 hours respectively,and a control group treated with DMEM complete medium only.Cell counting kit-8 (CCK8) assay was conducted to evaluate cellular proliferative activity to select the optimal concentration of trypsin.Then,some A431 cells treated with trypsin at the selected concentration for 24,48 and 72 hours respectively (or 48 hours only) served as the experimental groups (or group),and other A431 cells treated with DMEM complete medium served as the control group.Flow cytometry was performed to assess cell cycle distribution and proliferation index,fibronectin-based adhesion assay to estimate cell adhesive capacity,and wound healing assay and Transwell assay were conducted to evaluate the migratory capacity of cells in two-and three-dimensional space.Statistical analysis was carried out by using analysis of variance,paired samples t test and chi-square test.Results The proliferative activity of A431 cells increased along with the increase of trypsin concentrations,with the strongest increasing effect observed at 100 nmol/L.After treatment with 100 nmol/L trypsin,the experimental group showed a decrease in the percentage of G1-phase cells,but an increase in the percentage of S-phase cells,proliferation index,migratory and adhesive capacity compared with the control group (all P < 0.05).Conclusion Trypsin can promote the proliferation,migration and adhesion of A431 cells.
5.Quantification of Simian Immunodeficiency Virus by SYBR Green RT-PCR Technique
Jing LU ; Li QIN ; Guangjie LIU ; Siting ZHAO ; Xiaoping CHEN
Virologica Sinica 2008;23(3):189-195
Plasma viral RNA load is widely accepted as the most relevant parameter to assess the status and progression of Simian immunodeficiency virus (SIV) infections. To accurately measure RNA levels of the virus, a one-step fluorescent quantitative assay was established based on the SYBR green Real-time reverse transcription-polymerase chain reaction (RT-PCR). The lower detection limit of the assay was 10 copies per reaction for the virus. This method was successfully applied to quantify SIVmac251 and SIVmac239 viruses produced in CEM×174 cells. Additionally, the performance of the SYBR green RT-PCR was assessed in a SIVmac251 infected rhesus macaque. The result demonstrated that the method could detect as little as 215 copies per milliliter of plasma and the dynamic pattern of viral load was highly consistent with previous results. With regard to convenience, sensitivity and accuracy our assay represents a realistic alternative to both branched-chain DNA (b-DNA) assays or real-time PCR assays based on TaqMan probes.
6.Effects of morphine on PTEN expression and NF-κB activity in human gastric carcinoma cell line MGC-803
Yi QIN ; Jing CHEN ; Zhiling XIAO ; Yubo XIE ; Qiang XIAO
Chinese Journal of Anesthesiology 2010;30(12):1446-1448
Objective To investigate the effects of morphine on PTEN expression and NF-κB activity in human gastric carcinoma cell line MGC-803.Methods The human gastric cancer cell line MGC-803 was purchased from Cell Biology Research Institute,Chinese Academy of Sciences,and cultured in DMEM liquid culture medium.The cells were randomly divided into 2 groups(n = 6 each): control group and morphine group.The cells was exposed to 0.1 μmol/L morphine in morphine group.The apoptosis was assessed by flow cytometry after being incubated with morphine for 24 h.PTEN expression and NF-κB activity were detected using RT-PCR and Western blot.Results The apoptotic rate was significantly increased,PTEN expression was up-regulated and NF-κB activity was significantly decreased in morphine group compared with control group(P < 0.05).Conclusion Morphine can promote the apoptosis in human gastric cancer cells by up-regulating PTEN expression and decreasing NF-κB activity.
7.Relationship between C677T polymorphism of methylenetetrahydrofolate reductase gene/G448A polymorphism of β-fibrinogen gene and genetic susceptibility of ischemic stroke
Mulin HU ; Tao LI ; Qin CHEN ; Zhiwei JING ; Zhong WANG
International Journal of Cerebrovascular Diseases 2009;17(3):181-185
Objective To investigate the relationship between the C677T polymor-phism of the methylenetetrahydrofolate reductase (MTHFR) gene/G448A polymorphism of the
8.Effect of fentanyl on viability of human gastric carcinoma cell line MGC-803
Yi QIN ; Li LI ; Jing CHEN ; Yubo XIE
Chinese Journal of Anesthesiology 2010;30(6):705-707
Objective To investigate the effect of fentanyl on the viability of human gastric cancer cell line MGC-803. Methods The human gastric cancer cell line MGC-803 was purchased from Cell Biology Research Institute, Chinese Academy of Sciences, and cultured in DMEM liquid culture medium. The cells were seeded in 6-well or 96-well plates and divided into 3 groups (n = 60 wells each): group Ⅰ normal control (group C); group Ⅱ and Ⅲ were exposed to fentanyl 0.01 and 1.00 μmol/L respectively (group F1, F2). The viability of the cells was detected by MTT assay after being incubated with fentanyl for 12, 24, 36, 48, 60 and 72 h. The cell cycle progression and apoptosis were assessed by flow cytometry and the ulrastructure of the cells was examined with transmission electron microscope after being incubated with fentanyl for 24 h. The proliferation of the cells was determined by colony formation assay at 7 day of incubation with fentanyl. Results The viability and proliferation of the cells and the proportion of the cells in S phase were significantly lower, while the proportion of the cella in G2/M phase and the apoptotic rate were higher in group F1 and F2 than in group C but no significant difference was found between group F1 and F2. The nuclear evelope was intact, the nucleolus and chromosomes were clearly visible in group C, while in group F1 and F2 fregmentation of nuclear envelope and nucleolus, chromatin condensation and apoptotic bodies were observed in group F2. Conclusion Fentanyl can inhibit the viability of human gastric cancer cells by its pro-apoptosis inducing effect.
10.Nitric oxide and endoplasmic reticulum stress
Jing CHEN ; Jiong QIN ; Ying HAN ; Xiaoyan LIU
Chinese Journal of Pathophysiology 2000;0(11):-
Nitric oxide (NO) is a multifunctional biomolecule involved in a variety of physiological and pathological processes, including regulation of blood vessel dilatation and function as a neurotransmitter. However, a large amount of NO is toxic to the host and causes several diseases such as cardiovascular system diseases, septic shock, and diabetes mellitus. Endoplasmic reticulum (ER) stress pathway was first identified as a cellular response pathway induced by the accumulation of unfolded proteins in ER to preserve ER functions. Later it was found that ER stress pathway is also activated by various cellular stresses to protect cells, but when stresses are severe, apoptosis is induced to remove damaged cells. It is reported that NO disturbs ER functions, then ER stress-mediated apoptosis pathway is activated. CHOP/GADD153, which belongs to C/EBP transcription factor family, is induced in this process and mediates apoptosis. ER stress pathway induced by NO is involved in the pathogenesis of various diseases.